Type III collagen is found in most connective tissues, but is not found in hyaline cartilage. However, a unique type III collagen-related RNA has been detected in chick cartilage. This 3800 base RNA is much smaller than the authentic 5700 nucleotide type III collagen mRNA, and is in fact too small to encode a normal type III collagen subunit. Exons 21-52 of this RNA are similar or identical to the authentic type III collagen mRNA, but exons 1-16 appear to be missing from the 5' end of the RNA, suggesting that there may be an alternative transcription start site in the type III collagen gene which is used preferentially in cartilage. The absence of type III collagen in cartilage is presumably due to the altered RNA structure. Experiments are proposed to determine the structure of the type III collagen-related RNA and compare it with the authentic type III collagen mRNA. The 5' end of the chick type III collagen gene will also be isolated, and the promoter(s) and any previously undescribed exons will be identified by comparison of the exon/intron organization with the sequences of both the authentic type III collagen mRNA and the type III collagen-related RNA. These experiments will indicate whether the structural differences in the cartilage RNA are due to use of an alternative promoter, alternative RNA processing events, or both. They will also define any open reading frames in the RNA, thus indicating whether the RNA is likely to encode a protein (either a non-collagen protein or an altered form of type III collagen). The cis-acting sequences and trans-acting factors that are important for the function of the promoter(s) of the type III collagen gene in the appropriate tissues will be identified. Finally, the function of the cartilage type III collagen-related RNA will be determined. If, based on sequence and subcellular distribution data, the type III collagen-related RNA appears to encode a protein, the protein will be isolated and characterized. These experiments will provide increased understanding of the molecular mechanisms preventing production of type III collagen in cartilage, and will provide more extensive analysis of the structure of the type III collagen gene and its regulation than has heretofore been available. These studies on type III collagen gene expression, together with ongoing studies of alpha2(I) collagen gene expression, suggest the intriguing possibility that there may be a subset of collagen genes that encode alternative forms of mRNA in cartilage.